T-cell acute lymphoblastic leukemia (T-ALL) comprises about 15% of all cases of acute lymphoblastic leukemia (ALL). Features that distinguish this disease from B-lineage ALL include higher incidence in males, older mean age at diagnosis, high mean blood leukocyte count and the frequent presence of a mediastinal mass (Sen, L., and L. Borella (1975) "Clinical importance of lymphoblasts with T-markers in childhood leukemia," N. Engl. J. Med. 292:828-832; Tsukimoto, I., et al. (1976) "Surface markers and prognostic factors in acute lymphoblastic leukemia," N. Engl. J. Med. 294:245-248; Roper, M., et al. (1983) "Monoclonal antibody characterization of surface antigens in childhood T-cell lymphoid malignancies," Blood 61:830-837). Although significant improvements in long-term disease-free survival for both children (Shuster, J. J., et al. (1990) "Prognostic factors in childhood T-cell acute lymphoblastic leukemia: a Pediatric Oncology Group study," Blood 75:166-173; Pui, C. H., et al. (1990) "Heterogeneity of presenting features and their relation to treatment outcome in 120 children with T-cell acute lymphoblastic leukemia," Blood 75:174-179.) and adults (Hoelzer, D., et al. (1988) "Prognostic factors in a multicenter study for treatment of acute lymphoblastic leukemia in adults," Blood 71:123-131; Gaynor, J., et al. (1988) "A cause-specific hazard rate analysis of prognostic factors among 199 adults with acute lymphoblastic leukemia: The Memorial Hospital experience since 1969," J. Clin. Oncol. 6:1014-1030) have resulted from contemporary therapeutic programs, about 40% of children and at least 60% of adults with T-ALL still relapse and die of drug-resistant disease. Such failure is presumably due to residual leukemic cells which resist standard therapy. Therefore, a major challenge is to improve the detection and monitoring of minimal residual disease in patients receiving treatment. A sensitive and specific assay for residual leukemia is needed to direct the development of strategies for prevention of disease recurrence.
Since induction of complete remission is achieved in the vast majority of patients, current efforts to prevent treatment failure focus upon modifications of post-remission consolidation and/or maintenance chemotherapy. Unfortunately, the disease is not detectable by routine analysis during the remission period; thus, the effect of therapy on tumor burden is difficult to assess. A sensitive clonal assay for the residual leukemic population during remission would help guide therapeutic decisions beyond the induction period.
A number of chromosomal translocations are associated with T-ALL (Raimondi, S. C., et al. (1988) "Cytogenetics of childhood T-cell leukemia," Blood 72:1560-1566; Kaneko, Y., et al. (1989) "Chromosomal and immunophenotypic patterns in T cell acute lymphoblastic leukemia (T ALL) and lymphoblastic lymphoma (LBL)," Leukemia 3:886-892; Ucken, F. M., et al. (1989) "Immunophenotype-karyotype associations in human acute lymphoblastic leukemia," Blood 73:271-280). Theoretically, the underlying molecular rearrangements could form the basis for sensitive clonal assays for minimal residual disease (Delfau, M. H., et al. (1990) "Detection of minimal residual disease in chronic myeloid leukemia patients after bone marrow transplantation by polymerase chain reaction," Leukemia 4:1-5; Snyder, D. S., et al. (1989) "Definition of remission based on the expression of bcr-abl RNA following bone marrow transplant for chronic myelogenous leukemia in chronic phase," Blood 74 (Supl. 1):29a (Abstr.); Kohler, S., et al. (1989) "Application of the polymerase chain reaction to the detection of minimal residual disease after bone marrow transplantation for patients with chronic myelogenous leukemia," Blood 74 (Supl. 1):29a (Abstr.)). However, no single translocation is found in more than 5-10% of cases, and 20-30% of these leukemias display no karyotypic abnormalities at all (Raimondi, S. C., et al. (1988) "Cytogenetics of childhood T-cell leukemia," Blood 72:1560-1566; Kaneko, Y., et al. (1989) "Chromosomal and immunophenotypic patterns in T cell acute lymphoblastic leukemia (T ALL) and lymphoblastic lymphoma (LBL)," Leukemia 3:886-892; Ucken, F. M., et al. (1989) "Immunophenotype-karyotype associations in human acute lymphoblastic leukemia," Blood 73:271-280). This cytogenetic heterogeneity suggests that a number of loci will need to be characterized in detail before specific molecular probes suitable for detection of occult leukemia in the majority of cases can be prepared.
Chromosome 1 harbors a genetic locus (designated tal, for T-cell acute leukemia) involved in leukemogenesis. The tal-1 gene was identified upon analysis of t(1:14)(p34;q11). Recently, the breakpoint regions derived from one recurrent cytogenetic defect in T-ALL, namely the t(1;14)(p34;q11) translocation were isolated and sequenced (Chen, Q., et al. (1990) "The tal gene undergoes chromosome translocation in T cell leukemia and potentially encodes a helix-loop-helix protein," EMBO J. 9:415-424; Chen, Q., et al. (1990) "Coding sequences of the tal-1 gene are disrupted by chromosome translocation in human T cell leukemia," J. Exp. Med. 172:1403-1408). In 6 cases analyzed in detail, the breakpoints on chromosome 1 clustered within a 1 kb region. This translocation cleaves the tal-1 gene on chromosome 1, separating its 5' end from the rest of the gene which is transposed into the T cell receptor .alpha./.gamma. locus on chromosome 14. The tal-1 gene potentially encodes a protein containing a helix-loop-helix domain, which is found in a growing number of highly conserved DNA binding proteins involved in the regulation of growth and development (Murre, C., et al. (1989) "A new DNA binding and dimerization motif in immunoglobulin enhancer binding, daughterless, MyoD, and myc proteins," Cell 56:777-783). Several genes in this family are known to be disrupted in subsets of ALL (Leder, P., et al. (1983) "Translocations among antibody genes in human cancer," Science 222:765-771; Mellentin, J. D., et al. (1989) "lyl-1, a novel gene altered by chromosomal translocation in T cell leukemia, codes for a protein with a helix-loop-helix DNA binding motif," Cell 58:77-83; Mellentin, J. D., et al. (1989) "The gene for enhancer binding proteins E12/E47 lies at the t(1;19) breakpoint in acute leukemias," Science 246:379-382; Nourse, J., et al. (1990) "Chromosomal translocation t(1;19) results in synthesis of a homeobox fusion mRNA that codes for a potential chimeric transcription factor," Cell 60:535-545; Kamps, M. P., et al. (1990) "A new homeobox gene contributes the DNA binding domain of the t(1;19) translocation protein in pre-B ALL," Cell 60:547-555). Although of potential pathogenic significance, the t(1;14)(p34;q11) translocation is found in only 3% of T-ALLs (Carroll, A., et al. (1990) "The t(1;14)(p34;q11) translocation is non-random and restricted to T-cell acute lymphoblastic leukemia," Blood 76:1220-1224).
To investigate the possibility of a wider role for the tal-1 gene in T-ALL, rearrangements of this gene in the blast cells from a group of 50 patients with T-ALL were studied. These leukemias did not harbor the t(1;14)(p34;q11) translocation. Surprisingly, 13 (26%) of these leukemias contained rearrangements at this locus, all of which were identical at the level of Southern hybridization analysis (U.S. application Ser. No. 07/590,408, filed Sep. 28, 1990, now abandoned; Brown, L., et al. (1990) "Site-specific recombination of the tal-1 gene is a common occurrence in human T cell leukemia," EMBO J. 9:3343-3351). Detailed analysis revealed a site-specific of approximately 90 kb deletion on chromosome 1, one end of which lies about 1 kb from the clustered translocation breakpoints in the t(1;14)(p34;q11) cases. As demonstrated by nucleotide sequence analysis, the deletions are all remarkably site-specific, differing at their ends by only a few bases from one leukemia to another. These deletions were not found in remission peripheral blood leukocytes and therefore appear to be leukemia-specific. Thus, site-specific rearrangements at the tal-1 locus characterize nearly 30% of T-ALLs: about 3% are due to translocation (tal.sup.t alleles), while 26% result from an interstitial deletion which is too small to be detected cytogenetically (tal.sup.d alleles).
These rearrangements provide the opportunity to develop sensitive clonal assays for the relevant leukemias. In this invention, it is described assays which can detect 10 rearranged tal-1 cells in a background of 10.sup.6 normal cells. Moreover, a modification of the assay is presented which quantitates tal.sup.d alleles.
It is, therefore, desirable to have a sensitive assay, using a specific marker, for detecting and monitoring minimal residual leukemia cells in patients under treatment to develop strategies for the prevention of the recurrence of the disease.